Src Homology 3-interacting Domain of Rv1917c of Mycobacterium tuberculosis Induces Selective Maturation of Human Dendritic Cells by Regulating PI3K-MAPK-NF-kappa B Signaling and Drives Th2 Immune Responses

Mycobacterium tuberculosis, an etiological agent of pulmonary tuberculosis, causes significant morbidity and mortality worldwide. Pathogenic mycobacteria survive in the host by subverting host innate immunity. Dendritic cells (DCs) are professional antigen-presenting cells that are vital for eliciting immune responses to infectious agents, including pathogenic mycobacteria. DCs orchestrate distinct Th responses based on the signals they receive. In this perspective, deciphering the interactions of the proline-glutamic acid/proline-proline-glutamic acid (PE/PPE) family of proteins of M. tuberculosis with DCs assumes significant pathophysiological attributes. In this study, we demonstrate that Rv1917c (PPE34), a representative member of the proline-proline-glutamic-major polymorphic tandem repeat family, interacts with TLR2 and triggers functional maturation of human DCs. Signaling perturbations implicated a critical role for integrated cross-talk among PI3K-MAPK and NF-kappa B signaling cascades in Rv1917c-induced maturation of DCs. However, this maturation of DCs was associated with a secretion of high amounts of anti-inflammatory cytokine IL-10, whereas Th1-polarizing cytokine IL-12 was not induced. Consistent with these results, Rv1917c-matured DCs favored secretion of IL-4, IL-5, and IL-10 from CD4(+) T cells and contributed to Th2-skewed cytokine balance ex vivo in healthy individuals and in patients with pulmonary tuberculosis. Interestingly, the Rv1917c-skewed Th2 immune response involved induced expression of cyclooxygenase-2 (COX-2) in DCs. Taken together, these results indicate that Rv1917c facilitates a shift in the ensuing immunity toward the Th2 phenotype and could aid in immune evasion by mycobacteria.

A heuristic clustering algorithm using union of overlapping pattern-cells

Relative geometric arrangements of the sample points, with reference to the structure of the imbedding space, produce clusters. Hence, if each sample point is imagined to acquire a volume of a small M-cube (called pattern-cell), depending on the ranges of its (M) features and number (N) of samples; then overlapping pattern-cells would indicate naturally closer sample-points. A chain or blob of such overlapping cells would mean a cluster and separate clusters would not share a common pattern-cell between them. The conditions and an analytic method to find such an overlap are developed. A simple, intuitive, nonparametric clustering procedure, based on such overlapping pattern-cells is presented. It may be classified as an agglomerative, hierarchical, linkage-type clustering procedure. The algorithm is fast, requires low storage and can identify irregular clusters. Two extensions of the algorithm, to separate overlapping clusters and to estimate the nature of pattern distributions in the sample space, are also indicated.

A Comparative Study of the Effect of Metallic Au and ReO3 Nanoparticles on the Performance of Silicon Solar Cells

The influence of gold (similar to 35 nm diameter) as well as ReO3 (similar to 17 nm diameter) nanoparticles placed atop silicon photovoltaic devices on absorption and photocurrent generation has been investigated. The nanoparticles improve the power transmission into the semiconductor and consequently, the photocurrent response at wavelengths corresponding to plasmon absorption. An increase in short circuit current up to 4.5% under simulated solar irradiation was observed with the ReO3 nanoparticles, while the gold nanoparticles showed enhancements up to 6.5%. The increase in photocurrent is observed at wavelengths corresponding to the maxima in the surface plasmon resonance absorption spectra. (C) 2010 The Japan Society of Applied Physics

Photo-activated cytotoxicity of a pyrenyl-terpyridine copper(II) complex in HeLa cells

Terpyridine copper(II) complexes Cu(L)(2)](NO3)(2) where L is (4'-phenyl)-2 2' 6' 2 `'-terpyridine (ph-tpy in 1) and 4-(1 pyrenyl)]-2 2' 6' 2'-terpyridine (py-tpy in 2) are prepared characterized and their photocytotoxic activity studied The crystal structure of complex 1 shows distorted octahedral CuN6 coordination geometry The 1 2 electrolytic and one-electron paramagnetic complexes show a visible band near 650 nm in DMF-H2O The complexes show emission band at 352 nm for 1 and 425 nm for 2 when excited at 283 and 346 nm respectively The Cu(II)-Cu(I) redox couple is observed near -0 2 V versus SCE in DMF-0 1 m TBAP The complexes are avid partial-intercalative binders to calf thymus DNA giving binding constant (K-b) values of similar to 10(6) M-1 Complex 2 with its photoactive pyrenyl moiety exhibits significant photocleavage of pUC19 DNA in red light via singlet oxygen pathway Complex 2 also exhibits significant photo-activated cytotoxicity in HeLa cancer cells in visible light giving IC50 value of 11 9 mu M while being non-toxic in dark with an IC50 value of 130 5 mu M (C) 2010 Elsevier Ltd All rights reserved

Mesoporous Carbon and Poly(3,4-ethylenedioxythiophene) Composite as Catalyst Support for Polymer Electrolyte Fuel Cells

In situ polymerization of 3,4-ethylenedioxythiophene with sol-gel-derived mesoporous carbon (MC) leading to a new composite and its subsequent impregnation with Pt nanoparticles for application in polymer electrolyte fuel cells (PEFCs) is reported. The composite exhibits good dispersion and utilization of platinum nanoparticles akin to other commonly used microporous carbon materials, such as carbon black. Pt-supported MC-poly(3,4-ethylenedioxythiophene) (PEDOT) composite also exhibits promising electrocatalytic activity toward oxygen reduction reaction, which is central to PEFCs. The PEFC with Pt-loaded MC-PEDOT support exhibits 75% of enhancement in its power density in relation to the PEFC with Pt-loaded pristine MC support while operating under identical conditions. It is conjectured that Pt-supported MC-PEDOT composite ameliorates PEFC performance/durability on repetitive potential cycling. (C) 2010 The Electrochemical Society. DOI: 10.1149/1.3486172] All rights reserved.

Signaling different pathways of cell death: Abrin induced programmed necrosis in U266B1 cells

Abrin is a type II ribosome-inactivating protein comprising of two subunits, A and B. Of the two, the A-subunit harbours the RNA-N-glycosidase activity and the B subunit is a galactose specific lectin that enables the entry of the protein inside the cell. Abrin inhibits protein synthesis and has been reported to induce apoptosis in several cell types. Based on these observations abrin is considered to have potential for the construction of immunotoxin in cell targeted therapy. Preliminary data from our laboratory however showed that although abrin inhibited the protein synthesis in all cell types, the mode of cell death varied. The aim of the present study was therefore to understand different death pathways induced by abrin in different cells. We used the human B cell line, U266B1 and compared it with the earlier studied T cell line Jurkat, for abrin-mediated inhibition of protein translation as well as cell death. While abrin triggered programmed apoptosis in Jurkat cells in a caspase-dependent manner, it induced programmed necrosis in U266B1 cells in a caspase-independent manner, even when there was reactive oxygen species production and loss of mitochondrial membrane potential. The data revealed that abrin-mediated necrosis involves lysosomal membrane permeabilization and release of cathepsins from the lysosomes. Importantly, the choice of abrin-mediated death pathway in the cells appears to depend on which of the two events occurs first: lysosomal membrane permeabilization or loss of mitochondrial membrane potential that decides cell death by necrosis or apoptosis. (C) 2010 Elsevier Ltd. All rights reserved.

Temperature-Sensitive Src-Transformed Mdck Cells nn 3d Cultures as a Model of Malignant Transformation of Epithelial Cells

The equilibrium between cell proliferation, differentiation, and apoptosis is crucial for maintaining homeostasis in epithelial tissues. In order for the epithelium to function properly, individual cells must gain normal structural and functional polarity. The junctional proteins have an important role both in binding the cells together and in taking part in cell signaling. Cadherins form adherens junctions. Cadherins initiate the polarization process by first recognizing and binding the neighboring cells together, and then guiding the formation of tight junctions. Tight junctions form a barrier in dividing the plasma membranes to apical and basolateral membrane domains. In glandular tissues, single layered and polarized epithelium is folded into tubes or spheres, in which the basal side of the epithelial layer faces the outer basal membrane, and the apical side the lumen. In carcinogenesis, the differentiated architecture of an epithelial layer is disrupted. Filling of the luminal space is a hallmark of early epithelial tumors in tubular and glandular structures. In order for the transformed tumor cells to populate the lumen, enhanced proliferation as well as inhibition of apoptosis is required. Most advances in cancer biology have been achieved by using two-dimensional (2D) cell culture models, in which the cells are cultured on flat surfaces as monolayers. However, the 2D cultures are limited in their capacity to recapitulate the structural and functional features of tubular structures and to represent cell growth and differentiation in vivo. The development of three-dimensional (3D) cell culture methods enables the cells to grow and to be studied in a more natural environment. Despite the wide use of 2D cell culture models and the development of novel 3D culture methods, it is not clear how the change of the dimensionality of culture conditions alters the polarization and transformation process and the molecular mechanisms behind them. Src is a well-known oncogene. It is found in focal and adherens junctions of cultured cells. Active src disrupts cell-cell junctions and interferes with cell-matrix binding. It promotes cell motility and survival. Src transformation in 2D disrupts adherens junctions and the fibroblastic phenotype of the cells. In 3D, the adherens junctions are weakened, and in glandular structures, the lumen is filled with nonpolarized vital cells. Madin-Darby canine kidney (MDCK) cells are an epithelial cell type commonly used as a model for cell polarization. Its-src-transformed variants are useful model systems for analyzing the changes in cell morphology, and they play a role in src-induced malignant transformation. This study investigates src-transformed cells in 3D cell cultures as a model for malignant transformation. The following questions were posed. Firstly: What is the role of the composition and stiffness of the extracellular matrix (ECM) on the polarization and transformation of ts v-src MDCK cells in 3D cell cultures? Secondly: How do the culture conditions affect gene expression? What is the effect of v-src transformation in 2D and in 3D cell models? How does the shift from 2D to 3D affect cell polarity and gene expression? Thirdly: What is the role of survivin and its regulator phosphatase and tensin homolog protein (PTEN) in cell polarization and transformation, and in determining cell fate? How does their expression correlate with impaired mitochondrial function in transformed cells? In order to answer the above questions, novel methods of culturing and monitoring cells had to be created: novel 3D methods of culturing epithelial cells were engineered, enabling real time monitoring of a polarization and transformation process, and functional testing of 3D cell cultures. Novel 3D cell culture models and imaging techniques were created for the study. Attention was focused especially on confocal microscopy and live-cell imaging. Src-transformation disturbed the polarization of the epithelium by disrupting cell adhesion, and sensitized the cells to their environment. With active src, the morphology of the cell cluster depended on the composition and stiffness of the matrix. Gene expression studies revealed a broader impact of src transformation than mere continuous activity of src-kinase. In 2D cultures, src transformation altered the expression of immunological, actin cytoskeleton and extracellular matrix (ECM). In 3D, the genes regulating cell division, inhibition of apoptosis, cell metabolism, mitochondrial function, actin cytoskeleton and mechano-sensing proteins were altered. Surprisingly, changing the culture conditions from 2D to 3D affected also gene expression considerably. The microarray hit survivin, an inhibitor of apoptosis, played a crucial role in the survival and proliferation of src-transformed cells.

Atmospheric boundary-layer across Hadley and Ferrel cells over the Indian Ocean

A Southern Ocean Pilot cruise covering the latitudes from 10 degrees N to 56 degrees S in the open Indian Ocean was carried out during January February 2004. Surface and upper air data collected during this cruise are reported here. It is shown that the broad features of the atmosphere, in particular that of temperature, follow the tropical and mid-latitude weather expected during January February in this region. However, the atmospheric boundary-layer shows large variations, both in its height and structure between tropics and high latitudes. Strong influence of the surface heat flux on boundary layer structure is clearly seen. Humidity field reveals several local maxima and minima, suggesting a laminated atmosphere with air from different sources moving almost unmixed in adjacent layers.

Effect of 80 kDa protein of porcine follicular fluid on gonadotropin stimulated progesterone production in rat granulosa cells in vitro

We reported the presence of a 80 kDa polypeptide in porcine follicular fluid that inhibited the binding of 125I-radiolabelled hFSH as well as hCG to the rat ovarian gonadotropin receptors. In the present study, the biological activity of the receptor binding inhibitor is determined using an in vitro bioassay procedure. Granulosa cells isolated from PMSG primed immature rat ovaries respond to exogenously added gonadotropins in terms of progesterone production. Addition of fractions containing the gonadotropin receptor binding inhibitory activity inhibited progesterone production stimulated by the gonadotropins in a dose-dependent fashion. The receptor binding inhibitory activity was also capable of inhibiting progesterone production stimulated by PMSG, which has both FSH- and LH-like activities in rats. In contrast, progesterone production stimulated by dbcAMP was not inhibited by the receptor binding inhibitor. This result indicates that the site of action of the inhibitor is proximal to the formation of the cAMP. The above observations point out to a possible role for this factor in modulating gonadotropin activity at the ovarian level.

Drainage and separation factors for static foams containing agglomerates of microbial cells

The presence of cell agglomerates has been found to influence significantly the rates of liquid drainage from static foams. The process of drainage has been modelled by considering the foam to be made of pentagonal dodecahedral bubbles yielding films, nearly horizontal and nearly vertical Plateau borders. The films are assumed to drain into both kinds of Plateau borders equally. The horizontal Plateau borders are assumed to receive liquid from the films and drain into the vertical Plateau borders, which, in turn, form the main flow paths for gravity drainage. The drainage process is assumed to be similar to that for pure liquid until a stage is reached where the size of the cell agglomerates become equivalent to those of films and Plateau borders. Thereafter, a squeezing flow mechanism has been formulated where the aggromerates deform and flow. The model based on the above assumptions has been verified against experimental results and has been found to predict not only drainage data but also the separation of cell agglomerates from broths.

Metabolism and Translocation of Aminophospholipids in Mammalian Cells

In this study we investigated the metabolism, i.e. remodeling and translocation, of the aminophospholipids phosphatidylserine (PS) and phosphatidylethanolamine (PE). A new method for introduction of exogenous PS and PE molecular species to cultured cells was developed, and combined with mass spectrometry it enabled more detailed follow-up of the metabolism of single molecular species than previously. We found that I) exogenous PS and PE molecular species can be efficiently introduced to cultured cells without compromising cell integrity, II) PS and PE molecular species are remodeled by several phospholipases displaying selectivity based on phopholipid head group and acyl chain composition, III) PS decarboxylase (PSD) and Kennedy pathways provide a different PE molecular species composition to the cellular PE pool. In addition, PE species produced by these pathways are translocated from the site of synthesis to other cell compartments depending on their acyl chain composition. The data obtained in the present study helps to understand cellular phospholipid metabolism in more depth. The data show that effective labeling of cultured cells by exogenous phospholipids does not compromise cell viability and may be used to disturb cellular phospholipid composition to study lipid homeostasis. Remodeling and translocation of PS and PE molecular species is highly selective. The developed method and mass- spectrometric techniques may be used in future studies to understand disturbances in lipid homeostasis for example in diabetes mellitus, thus opening doors to optional scientific approaches to study mechanisms behind pathologies related to lipid disturbances.

Nafion®-bound carbon electrodes containing transitionmetal phthalocyanines for oxygen reduction in solid-polymer-electrolyte fuel cells

Catalytic activities of some transition metal-phthalocyanine complexes towards electroreduction of molecular oxygen are examined on Nafion®-bound and bare porous carbon electrodes in 2.5 M H2SO4 electrolyte. It is found that these metal complexes exhibit better catalytic activities towards oxygen reduction with the Nafion®-bound electrodes.

Translational Up-Regulation and High-Level Protein Expression from Plasmid Vectors by mTOR Activation via Different Pathways in PC3 and 293T Cells

Background: Though 293T cells are widely used for expression of proteins from transfected plasmid vectors, the molecular basis for the high-level expression is yet to be understood. We recently identified the prostate carcinoma cell line PC3 to be as efficient as 293T in protein expression. This study was undertaken to decipher the molecular basis of high-level expression in these two cell lines. Methodology/Principal Findings: In a survey of different cell lines for efficient expression of platelet-derived growth factor-B (PDGF-B), beta-galactosidase (beta-gal) and green fluorescent protein (GFP) from plasmid vectors, PC3 was found to express at 5-50-fold higher levels compared to the bone metastatic prostate carcinoma cell line PC3BM and many other cell lines. Further, the efficiency of transfection and level of expression of the reporters in PC3 were comparable to that in 293T. Comparative analyses revealed that the high level expression of the reporters in the two cell lines was due to increased translational efficiency. While phosphatidic acid (PA)-mediated activation of mTOR, as revealed by drastic reduction in reporter expression by n-butanol, primarily contributed to the high level expression in PC3, multiple pathways involving PA, PI3K/Akt and ERK1/2 appear to contribute to the abundant reporter expression in 293T. Thus the extent of translational upregulation attained through the concerted activation of mTOR by multiple pathways in 293T could be achieved through its activation primarily by the PA pathway in PC3. Conclusions/Significance: Our studies reveal that the high-level expression of proteins from plasmid vectors is effected by translational up-regulation through mTOR activation via different signaling pathways in the two cell lines and that PC3 is as efficient as 293T for recombinant protein expression. Further, PC3 offers an advantage in that the level of expression of the protein can be regulated by simple addition of n-butanol to the culture medium.

Taking Multiple Infections of Cells and Recombination into Account Leads to Small Within-Host Effective-Population-Size Estimates of HIV-1

Whether HIV-1 evolution in infected individuals is dominated by deterministic or stochastic effects remains unclear because current estimates of the effective population size of HIV-1 in vivo, N-e, are widely varying. Models assuming HIV-1 evolution to be neutral estimate N-e similar to 10(2)-10(4), smaller than the inverse mutation rate of HIV-1 (similar to 10(5)), implying the predominance of stochastic forces. In contrast, a model that includes selection estimates N-e>10(5), suggesting that deterministic forces would hold sway. The consequent uncertainty in the nature of HIV-1 evolution compromises our ability to describe disease progression and outcomes of therapy. We perform detailed bit-string simulations of viral evolution that consider large genome lengths and incorporate the key evolutionary processes underlying the genomic diversification of HIV-1 in infected individuals, namely, mutation, multiple infections of cells, recombination, selection, and epistatic interactions between multiple loci. Our simulations describe quantitatively the evolution of HIV-1 diversity and divergence in patients. From comparisons of our simulations with patient data, we estimate N-e similar to 10(3)-10(4), implying predominantly stochastic evolution. Interestingly, we find that N-e and the viral generation time are correlated with the disease progression time, presenting a route to a priori prediction of disease progression in patients. Further, we show that the previous estimate of N-e>10(5) reduces as the frequencies of multiple infections of cells and recombination assumed increase. Our simulations with N-e similar to 10(3)-10(4) may be employed to estimate markers of disease progression and outcomes of therapy that depend on the evolution of viral diversity and divergence.

Temperature dependence of hydrogen evolution reaction on nickel oxide electrode in sealed nickel/cadmium cells

The impedance of sealed nickel/cadmium cells around a cell e.m.f. of 0.0 V was measured at five different temperatures between � 10 and +30 °C. The results show that the behaviour is similar at all temperatures. Based on the experimental results, the relation between charge-transfer resistance (Rct) and temperature (T) has been established for the Volmer reaction. Further, the value of cathodic transfer coefficient (?) has been estimated.